The major goals of this research are to use genetic approaches to determine: (a) whether metallothioneins (MT) are essential during mammalian development and physiology, (b) whether different isoforms of MT have distinct functions and (c) whether MTs can play a role in retarding malignant transformation. The MT-I and MT-II genes are expressed in most organs, they are inducible by a variety of metals and hormones, and they probably have a similar, general function. These MTs will be disrupted by homologous recombination in embryonic stem cells and those cells will be used to generate mice lacking MT-I, MT-II or both isoforms. The effect of these mutations on normal development and physiological responses will be examined. Mice over-expressing MT-I or MT-III (an unusual MT normally expressed only in the brain) have been generated by microinjection of marked versions of these genes flanked by presumptive locus control regions from the MT locus. These transgenic mice will be used to determine if excess or ectopic expression of MT affects development or physiology. Mice expressing reduced or elevated levels of MT will then be crossed with two different transgenic lines of mice with predispositions to develop hepatocellular carcinoma. If MT plays a role in retarding tumorigenesis, then we predict that hepatic tumors will develop more slowly in mice expressing excess MT and more rapidly in mice expressing reduced levels of MT.